The non-histone proteins of chromatin, their isolation and composition in a number of tissues.

Abstract

1. 1. A method is described for the fractionation of salt-urea-dissociated chromatin using hydroxylapatite. With the exception of experiments using chromatins prepared from “citric acid” nuclei, high yields of acidic non-histone proteins, relatively free of RNA, can be obtained by this procedure. 2. 2. The non-histone proteins of a number of chromatins were compared by electrophoresis in sodium dodecyl sulphate-urea polyacrylamide gels employing a discontinuous buffer system. Proteins from mouse chromatins prepared from “citric acid” nuclei were found to be extremely heterogeneous, but in the case of calf thymus the proteins were mainly low molecular weight. On the other hand, the non-histone proteins of chromatin from “sucrose” nuclei appeared to contain fewer high molecular weight species in the tissues studied, with the exception of brain. Preparation of nuclei by the double-detergent procedure of Penman ( J. Mol. Biol. , 17 (1966) 117) gave chromatin with a low protein to DNA ratio. These proteins also appeared to be predominantly low molecular weight. Duck erythrocyte nuclei prepared by lysis also contained low molecular weight chromatin non-histone proteins. 3. 3. Using salt fractionation techniques attempts were also made to remove “cytoplasmic” and “residual” acidic proteins from chromatin. The proteins which remained with the DNA and histones were found to be mainly low molecular weight in kidney and liver, but in the case of brain a wide spectrum of proteins was seen. 4. 4. Little tissue or species specificity of non-histone proteins were found on comparison of “sucrose” nuclei chromatins prepared from a number of mouse and bovine tissues. 5. 5. It is concluded that the non-histone proteins which remain tightly bound to DNA in chromatin are of the same approximate size as the basic histones. Because of the procedural variation in the heterogeneity of these chromatin proteins, detection of single species of regulatory proteins appears to require other techniques.